In conclusion over expression of Stokesia epoxygenase SlEPX

2019-09-11

In conclusion, over-expression of Stokesia epoxygenase (SlEPX) in soybean seeds led to some unusual seed phenotypes. These effects can be overcome by coexpression of Vernonia DGATs (VgDGAT1 & 2). The DGATs can specifically transfer vernolic WZB117 into TAG, largely reducing vernolic acid levels in membranes, thus stabilizing cellular metabolism.
Acknowledgments
This work was supported by the Consortium for Plant Biotechnology Research (CPBR), the United Soybean Board (USB), Ashland Chemicals and the Kentucky Agricultural Experiment Station. We thank Scott Serdoz, Dennis Egli and Todd Pfeiffer for help with the field production of the soybeans, John Johnson for fatty acid analysis and Bob Geneve for help with pictures.
Obesity, generally defined as a body mass index (BMI) of ≥30 kg/m, is a major health problem in the United States, affecting ∼20% of adults . It is a risk factor for hypertension, diabetes, and cardiovascular disease. Health care costs attributable to obesity amount to ∼$100 billion annually . Despite recent discoveries concerning the regulation of energy metabolism, little is known about the pathogenesis of obesity, and few effective therapies exist .
Obesity can be viewed as an energy storage disorder . Weight gain results from an energy imbalance (i.e., energy input exceeding output), with most excess calories stored as triglycerides in adipose tissue. Over the past decade, genes and molecules that regulate energy intake and output (e.g., leptin and its receptor, neuropeptide Y, molecules in the melanocortin pathway) have been identified and studied extensively , . These discoveries have led to the identification of several metabolic pathways involved in energy regulation. However, the relationship between triglyceride synthesis and obesity has not been well studied. Specifically, it has been unclear whether disrupting triglyceride synthesis would be feasible.
Role of DGAT in Triglyceride Synthesis
A key enzyme in triglyceride synthesis is acyl CoA:diacylglycerol acyltransferase (DGAT), a microsomal enzyme that is widely expressed in mammalian tissues (reviewed in Farese et al. 2000). DGAT catalyzes the final reaction in the glycerol phosphate pathway, considered the main pathway of triglyceride synthesis in cells (Figure 2A). The enzyme is also believed to catalyze the final step of the monoacylglycerol pathway, found predominantly in enterocytes of the small intestine. In both pathways, DGAT facilitates the joining of a diacylglycerol with a fatty acyl CoA, resulting in the formation of triglyceride (Figure 2B). Although it is unclear whether DGAT is rate-limiting for triglyceride synthesis (Lehner and Kuksis 1996), it catalyzes the only step in the pathway that is committed to producing this type of compound.
While the biochemistry of DGAT has been studied for over 40 years, the DGAT gene had not been cloned because the enzyme\'s membrane-bound nature made its purification difficult. In 1998, we identified the gene from sequence database searches because of its similarity to acyl CoA:cholesterol acyltransferase (ACAT) genes (Cases et al. 1998). Homologs have been identified in both plant and animal species Bouvier-Navé et al. 2000, Hobbs et al. 1999, Oelkers et al. 1998. The cloning of DGAT has allowed molecular studies of DGAT to be performed for the first time. Northern blot analyses of DGAT mRNA levels correlated with previous studies of DGAT enzymatic activity; namely, the enzyme is expressed in all tissues examined, with the highest expression levels in liver, small intestine, and adipose tissue Cases et al. 1998, Farese et al. 2000, Oelkers et al. 1998. Significant DGAT expression is also detected in skeletal muscle and brain.
DGAT-Deficient Mice are Resistant to Obesity
To address questions concerning the function of DGAT, we inactivated its gene in mice (Smith et al. 2000). Surprisingly, DGAT knockout (Dgat−/−) mice, although deficient in DGAT activity, were viable and capable of synthesizing triglycerides, as evidenced by normal fasting serum triglyceride levels and normal adipose tissue composition. When fed a regular chow diet (4% fat by weight), the knockout mice exhibited weight curves similar to those of wild-type mice.